bims-mitdis 2021-12-19 papers

bims-mitdis

Biomed News

on Mitochondrial disorders

Issue of 2021–12–19
33 papers selected by
Catalina Vasilescu, University of Helsinki

Multifaceted Roles of Mitochondrial Stress Responses under ETC Dysfunction - Repair, Destruction and Pathogenesis.
Cristae junction as a fundamental switchboard for mitochondrial ion signaling and bioenergetics.
Genetically encoded biosensors for evaluating NAD+/NADH ratio in cytosolic and mitochondrial compartments.
The role of mitochondrial dynamics in mtDNA maintenance.
The mining and construction of a knowledge base for gene-disease association in mitochondrial diseases.
Dysfunctional mitochondrial translation and combined oxidative phosphorylation deficiency in a mouse model of hepatoencephalopathy due to Gfm1 mutations.
Cannabidiol activates PINK1-Parkin-dependent mitophagy and mitochondrial-derived vesicles.
Imaging of Mitophagy Enabled by an Acidity-Reporting Probe Anchored on the Mitochondrial Inner Membrane.
Molecular biomarkers correlate with brain grey and white matter changes in patients with mitochondrial m.3243A > G mutation.
Dysregulated mitochondrial and cytosolic tRNA m1A methylation in Alzheimer's disease.
MRE11-dependent instability in mitochondrial DNA fork protection activates a cGAS immune signaling pathway.
PINK1 signalling in neurodegenerative disease.
SSBP1-Disease Update: Expanding the Genetic and Clinical Spectrum, Reporting Variable Penetrance and Confirming Recessive Inheritance.
Rare presentation of FDX2-related disorder and untargeted global metabolomics findings.
Enhanced hepatic respiratory capacity and altered lipid metabolism support metabolic homeostasis during short-term hypoxic stress.
TRAP1 inhibits MIC60 ubiquitination to mitigate the injury of cardiomyocytes and protect mitochondria in extracellular acidosis.
Genetic landscape of human mitochondrial genome using whole genome sequencing.
Mutations in LRRK2 linked to Parkinson disease sequester Rab8a to damaged lysosomes and regulate transferrin-mediated iron uptake in microglia.
Mitochondrial hepatopathy: Respiratory chain disorders- 'breathing in and out of the liver'.
The molecular role of Sigmar1 in regulating mitochondrial function through mitochondrial localization in cardiomyocytes.
Reducing NAD(H) to amplify rhythms.
Circulating mitochondrial DNA copy numbers represent a sensitive marker for diagnosis and monitoring of disease activity in systemic lupus erythematosus.
Culturing Human Pluripotent Stem Cells on Micropatterned Silicon Surfaces.
What is the most appropriate covariate in ANCOVA when analysing metabolic rate?
Neurogenic Hypertension Mediated Mitochondrial Abnormality Leads to Cardiomyopathy: Contribution of UPRmt and Norepinephrine-miR- 18a-5p-HIF-1α Axis.
Reactive microglia and mitochondrial unfolded protein response following ventriculomegaly and behavior defects in kaolin-induced hydrocephalus.
Development of a Standardized MRM Method for the Quantification of One Carbon Metabolism Enzymes.
Murine cerebral organoids develop network of functional neurons and hippocampal brain region identity.
Altering neuronal circuitry with 4-aminopyridine for visual improvement in leber's hereditary optic neuropathy (LHON).
DNA methylation episignature testing improves molecular diagnosis of Mendelian chromatinopathies.
The importance of automation in genetic diagnosis: Lessons from analyzing an inherited retinal degeneration cohort with the Mendelian Analysis Toolkit (MATK).
Sarm1 haploinsufficiency or low expression levels after antisense oligonucleotides delay programmed axon degeneration.
Cost-effectiveness of genome sequencing for diagnosing patients with undiagnosed rare genetic diseases.

FEBS J. 2021 Dec 16.

Multifaceted Roles of Mitochondrial Stress Responses under ETC Dysfunction - Repair, Destruction and Pathogenesis.

Shanshan Liu, Siqi Liu, Hui Jiang.

  Electron transport chain (ETC) dysfunction is a common feature of mitochondrial diseases and induces severe cellular stresses, including mitochondrial membrane potential (Δψm ) reduction, mitochondrial matrix acidification, metabolic derangements and proteostatic stresses. Extensive studies of ETC dysfunction in yeast, C. elegans, cultured cells and mouse models have revealed multiple mitochondrial stress response pathways. Here, we summarize the current understanding of the triggers, sensors, signaling mechanisms, and the functional outcomes of mitochondrial stress responses in different species. We highlight Δψm reduction as a major trigger of stress responses in different species, but the responses are species-specific and the outcomes are context-dependent. ETC dysfunction elicits a mitochondrial unfolded protein response (UPRmt ) to repair damaged mitochondria in C. elegans, and activates a global adaptive program to maintain Δψm in yeast. Yeast and C. elegans responses are remarkably similar at the downstream responses, although they are activated by different signaling mechanisms. UPRmt generally protects ETC-defective worms, but its constitutive activation is toxic for wildtype worms and worms carrying mutant mtDNA. In contrast to lower organisms, ETC dysfunction in mammals mainly activates a mitochondrial integrated stress response (ISRmt ) to reprogram metabolism and a PINK1-Parkin mitophagy pathway to degrade damaged mitochondria. Accumulating in vivo results suggest that the ATF4 branch of ISRmt exacerbates metabolic derangements to accelerate mitochondrial disease progression. The in vivo roles of mitophagy in mitochondrial diseases are also context-dependent. These results thus reveal the common and unique aspects of mitochondrial stress responses in different species and highlight their multifaceted roles in mitochondrial diseases.

Keywords:  ISRmt; Mitochondrial stress response; UPRmt; mitochondrial membrane potential; mitophagy

DOI:  https://doi.org/10.1111/febs.16323
Cell Calcium. 2021 Dec 10. pii: S0143-4160(21)00171-8. [Epub ahead of print]101 102517

Cristae junction as a fundamental switchboard for mitochondrial ion signaling and bioenergetics.

Benjamin Gottschalk, Corina T Madreiter-Sokolowski, Wolfgang F Graier.

OPA1 and MICU1 are both involved in the regulation of mitochondrial Ca2+ uptake and the stabilization of the cristae junction, which separates the inner mitochondrial membrane into the interboundary membrane and the cristae membrane. In this mini-review, we focus on the synergetic control of OPA1 and MICU1 on the cristae junction that serves as a fundamental regulator of multiple mitochondrial functions. In particular, we point to the critical role of an adaptive cristae junction permeability in mitochondrial Ca2+ signaling, spatial H+ gradients and mitochondrial membrane potential, metabolic activity, and apoptosis. These characteristics bear on a distinct localization of the oxidative phosphorylation machinery, the FoF1-ATPase, and mitochondrial Ca2+uniporter (MCU) within sections of the inner mitochondrial membrane isolated by the cristae junction and regulated by proteins like OPA1 and MICU1. We specifically focus on the impact of MICU1-regulated cristae junction on the activity and distribution of MCU within the complex ultrastructure of mitochondria.

DOI: https://doi.org/10.1016/j.ceca.2021.102517
Cell Rep Methods. 2021 Nov 22. pii: 100116. [Epub ahead of print]1(7):

Genetically encoded biosensors for evaluating NAD+/NADH ratio in cytosolic and mitochondrial compartments.

Qingxun Hu, Dan Wu, Matthew Walker, Pei Wang, Rong Tian, Wang Wang.

The ratio of oxidized to reduced NAD (NAD+/NADH) sets intracellular redox balance and antioxidant capacity. Intracellular NAD is compartmentalized and the mitochondrial NAD+/NADH ratio is intricately linked to cellular function. Here, we report the monitoring of the NAD+/NADH ratio in mitochondrial and cytosolic compartments in live cells by using a modified genetic biosensor (SoNar). The fluorescence signal of SoNar targeted to mitochondria (mt-SoNar) or cytosol (ct-SoNar) responded linearly to physiological NAD+/NADH ratios in situ. NAD+/NADH ratios in cytosol versus mitochondria responded rapidly, but differently, to acute metabolic perturbations, indicating distinct NAD pools. Subcellular NAD redox balance regained homeostasis via communications through malate-aspartate shuttle. Mitochondrial and cytosolic NAD+/NADH ratios are influenced by NAD+ precursor levels and are distinctly regulated under pathophysiological conditions. Compartment-targeted biosensors and real-time imaging allow assessment of subcellular NAD+/NADH redox signaling in live cells, enabling future mechanistic research of NAD redox in cell biology and disease development.

DOI: https://doi.org/10.1016/j.crmeth.2021.100116
J Cell Sci. 2021 Dec 15. pii: jcs258944. [Epub ahead of print]134(24):

The role of mitochondrial dynamics in mtDNA maintenance.

Rasha Sabouny, Timothy E Shutt.

  The dynamic nature of mitochondria, which can fuse, divide and move throughout the cell, allows these critical organelles to adapt their function in response to cellular demands, and is also important for regulating mitochondrial DNA (mtDNA). While it is established that impairments in mitochondrial fusion and fission impact the mitochondrial genome and can lead to mtDNA depletion, abnormal nucleoid organization or accumulation of deletions, it is not entirely clear how or why remodeling mitochondrial network morphology affects mtDNA. Here, we focus on recent advances in our understanding of how mitochondrial dynamics contribute to the regulation of mtDNA and discuss links to human disease.

Keywords:  Fission; Fusion; Mitochondria; Mitochondrial dynamics; Mitophagy; mtDNA

DOI:  https://doi.org/10.1242/jcs.258944
Sci Rep. 2021 Dec 13. 11(1): 23909

The mining and construction of a knowledge base for gene-disease association in mitochondrial diseases.

Wei Wang, Junying Song, Yunhai Chuai, Fu Chen, Chunlan Song, Mingming Shu, Yayun Wang, Yunfei Li, Xinyu Zhai, Shujie Han, Shun Yao, Kexin Shen, Wei Shang, Lei Zhang.

Mitochondrial diseases are a group of heterogeneous genetic metabolic diseases caused by mitochondrial DNA (mtDNA) or nuclear DNA (nDNA) gene mutations. Mining the gene-disease association of mitochondrial diseases is helpful for understanding the pathogenesis of mitochondrial diseases, for carrying out early clinical diagnosis for related diseases, and for formulating better treatment strategies for mitochondrial diseases. This project researched the relationship between genes and mitochondrial diseases, combined the Malacards, Genecards, and MITOMAP disease databases to mine the knowledge on mitochondrial diseases and genes, used database integration and the sequencing method of the phenolyzer tool to integrate disease-related genes from different databases, and sorted the disease-related candidate genes. Finally, we screened 531 mitochondrial related diseases, extracted 26,723 genes directly or indirectly related to mitochondria, collected 24,602 variant sites on 1474 genes, and established a mitochondrial disease knowledge base (MitDisease) with a core of genes, diseases, and variants. This knowledge base is helpful for clinicians who want to combine the results of gene testing for diagnosis, to understand the occurrence and development of mitochondrial diseases, and to develop corresponding treatment methods.

DOI: https://doi.org/10.1038/s41598-021-03249-0
FASEB J. 2022 Jan;36(1): e22091

Dysfunctional mitochondrial translation and combined oxidative phosphorylation deficiency in a mouse model of hepatoencephalopathy due to Gfm1 mutations.

Miguel Molina-Berenguer, Ferran Vila-Julià, Sandra Pérez-Ramos, Maria Teresa Salcedo-Allende, Yolanda Cámara, Javier Torres-Torronteras, Ramon Martí.

  Hepatoencephalopathy due to combined oxidative phosphorylation deficiency type 1 (COXPD1) is a recessive mitochondrial translation disorder caused by mutations in GFM1, a nuclear gene encoding mitochondrial elongation factor G1 (EFG1). Patients with COXPD1 typically present hepatoencephalopathy early after birth with rapid disease progression, and usually die within the first few weeks or years of life. We have generated two different mouse models: a Gfm1 knock-in (KI) harboring the p.R671C missense mutation, found in at least 10 patients who survived more than 1 year, and a Gfm1 knock-out (KO) model. Homozygous KO mice (Gfm1-/- ) were embryonically lethal, whereas homozygous KI (Gfm1R671C / R671C ) mice were viable and showed normal growth. R671C mutation in Gfm1 caused drastic reductions in the mitochondrial EFG1 protein content in different organs. Six- to eight-week-old Gfm1R671C / R671C mice showed partial reductions of in organello mitochondrial translation and respiratory complex IV enzyme activity in the liver. Compound heterozygous Gfm1R671C /- showed a more pronounced decrease of EFG1 protein in liver and brain mitochondria, as compared with Gfm1R671C / R671C mice. At 8 weeks of age, their mitochondrial translation rates were significantly reduced in both tissues. Additionally, Gfm1R671C /- mice showed combined oxidative phosphorylation deficiency (reduced complex I and IV enzyme activities in liver and brain), and blue native polyacrylamide gel electrophoresis analysis revealed lower amounts of both affected complexes. We conclude that the compound heterozygous Gfm1R671C /- mouse presents a clear dysfunctional molecular phenotype, showing impaired mitochondrial translation and combined respiratory chain dysfunction, making it a suitable animal model for the study of COXPD1.

Keywords:   Gfm1 ; COXPD1; animal model; elongation factor G1; mitochondria; translation

DOI:  https://doi.org/10.1096/fj.202100819RRR
Eur J Cell Biol. 2021 Nov 14. pii: S0171-9335(21)00036-4. [Epub ahead of print]101(1): 151185

Cannabidiol activates PINK1-Parkin-dependent mitophagy and mitochondrial-derived vesicles.

Adrian Ramirez, William Old, David L Selwood, Xuedong Liu.

  The PINK1/Parkin pathway plays an important role in maintaining a healthy pool of mitochondria. Activation of this pathway can lead to apoptosis, mitophagy, or mitochondrial-derived vesicle formation, depending on the nature of mitochondrial damage. The signaling by which PINK/Parkin activation leads to these different mitochondrial outcomes remains understudied. Here we present evidence that cannabidiol (CBD) activates the PINK1-Parkin pathway in a unique manner. CBD stimulates PINK1-dependent Parkin mitochondrial recruitment similarly to other well-studied Parkin activators but with a distinctive shift in the temporal dynamics and mitochondrial fates. The mitochondrial permeability transition pore inhibitor cyclosporine A exclusively diminished the CBD-induced PINK1/Parkin activation and its associated mitochondrial effects. Unexpectedly, CBD treatment also induced elevated production of mitochondrial-derived vesicles (MDV), a potential quality control mechanism that may help repair partial damaged mitochondria. Our results suggest that CBD may engage the PINK1-Parkin pathway to produce MDV and repair mitochondrial lesions via mitochondrial permeability transition pore opening. This work uncovered a novel link between CBD and PINK1/Parkin-dependent MDV production in mitochondrial health regulation.

Keywords:  Cannabidiol; Mitochondrial quality control; Mitochondrial-derived vesicles (MDV); Mitophagy; PINK; Parkin

DOI:  https://doi.org/10.1016/j.ejcb.2021.151185
Anal Chem. 2021 Dec 12.

Imaging of Mitophagy Enabled by an Acidity-Reporting Probe Anchored on the Mitochondrial Inner Membrane.

Shixiong Wen, Xiao Hu, Yilong Shi, Jiahuai Han, Shoufa Han.

Classical chemical probes are prone to dissipation from stressed organelles, as evidenced by the incapability of mitochondrial dyes to image mitophagy linked to multiple diseases. We herein reported mitophagy imaging via covalent anchoring of a lysosomal probe to the mitochondrial inner membrane (CALM). Utilizing DBCORC-TPP, an azide-conjugatable probe with acidity-triggered fluorescence, CALM is operated via ΔΨm-promoted probe accumulation in mitochondria and thereby bioorthogonal ligation of the trapped probe with azido-choline (Azcholine) metabolically installed on the mitochondrial membrane. Overcoming the limitation of synthetic probes to dissipate from stressed organelles, CALM enables signal-on fluorescence imaging of mitophagy induced by starvation and is further employed to reveal mitophagy in ferroptosis. These results suggest the potential of CALM as a new tool to study mitophagy.

DOI: https://doi.org/10.1021/acs.analchem.1c03881
Mol Genet Metab. 2021 Nov 30. pii: S1096-7192(21)00830-1. [Epub ahead of print]

Molecular biomarkers correlate with brain grey and white matter changes in patients with mitochondrial m.3243A > G mutation.

Stefania Evangelisti, Laura Ludovica Gramegna, Chiara La Morgia, Lidia Di Vito, Alessandra Maresca, Lia Talozzi, Claudio Bianchini, Micaela Mitolo, David Neil Manners, Leonardo Caporali, Maria Lucia Valentino, Rocco Liguori, Valerio Carelli, Raffaele Lodi, Claudia Testa, Caterina Tonon.

   INTRODUCTION: The mitochondrial DNA (mtDNA) m.3243A > G mutation in the MT-TL1 gene results in a multi-systemic disease, that is commonly associated with neurodegenerative changes in the brain.
METHODS: Seventeen patients harboring the m3243A > G mutation were enrolled (age 43.1 ± 11.4 years, 10 M/7F). A panel of plasma biomarkers including lactate acid, alanine, L-arginine, fibroblast growth factor 21 (FGF-21), growth/differentiation factor 15 (GDF-15) and circulating cell free -mtDNA (ccf-mtDNA), as well as blood, urine and muscle mtDNA heteroplasmy were evaluated. Patients also underwent a brain standardized MR protocol that included volumetric T1-weighted images and diffusion-weighted MRI. Twenty sex- and age-matched healthy controls were included. Voxel-wise analysis was performed on T1-weighted and diffusion imaging, respectively with VBM (voxel-based morphometry) and TBSS (Tract-based Spatial Statistics). Ventricular lactate was also evaluated by 1H-MR spectroscopy.
RESULTS: A widespread cortical gray matter (GM) loss was observed, more severe (p < 0.001) in the bilateral calcarine, insular, frontal and parietal cortex, along with infratentorial cerebellar cortex. High urine mtDNA mutation load, high levels of plasma lactate and alanine, low levels of plasma arginine, high levels of serum FGF-21 and ventricular lactate accumulation significantly (p < 0.05) correlated with the reduced brain GM density. Widespread microstructural alterations were highlighted in the white matter, significantly (p < 0.05) correlated with plasma alanine and arginine levels, with mtDNA mutation load in urine, with high level of serum GDF-15 and with high content of plasma ccf-mtDNA.
CONCLUSIONS: Our results suggest that the synergy of two pathogenic mechanisms, mtDNA-related mitochondrial respiratory deficiency and defective nitric oxide metabolism, contributes to the brain neurodegeneration in m.3243A > G patients.

Keywords:  Brain gray matter atrophy; Brain white matter microstructure; M.3243A > G mutation; MR neuroimaging; Molecular biomarkers

DOI:  https://doi.org/10.1016/j.ymgme.2021.11.012
Hum Mol Genet. 2021 Dec 13. pii: ddab357. [Epub ahead of print]

Dysregulated mitochondrial and cytosolic tRNA m1A methylation in Alzheimer's disease.

Andrew M Shafik, Huiqing Zhou, Junghwa Lim, Bryan Dickinson, Peng Jin.

RNA modifications affect many aspects of RNA metabolism and are involved in the regulation of many different biological processes. Mono-methylation of adenosine in the N1 position, N1-methyladensoine (m1A), is a reversible modification that is known to target rRNAs and tRNAs. m1A has been shown to increase tRNA structural stability and induce correct tRNA folding. Recent studies have begun to associate the dysregulation of epitranscriptomic control with age-related disorders such as Alzheimer's disease. Here, we applied the newly developed m1A-quant-seq approach to map the brain abundant m1A RNA modification in the cortex of an Alzheimer's disease mouse model, 5XFAD. We observed hypomethylation in both mitochondrial and cytosolic tRNAs in 5XFAD mice compared to wild type. Furthermore, the main enzymes responsible for the addition of m1A in mitochondrial (TRMT10C, HSD17B10) and cytosolic tRNAs (TRMT61A) displayed decreased expression in 5XFAD compared to wild type mice. Knockdown of these enzymes results in a more severe phenotype in a Drosophila tau model, and differential m1A methylation is correlated with differences in mature mitochondrial tRNA expression. Collectively, this work suggests that hypo m1A modification in tRNAs may play a role in Alzheimer's disease pathogenesis.

DOI: https://doi.org/10.1093/hmg/ddab357
Sci Adv. 2021 Dec 17. 7(51): eabf9441

MRE11-dependent instability in mitochondrial DNA fork protection activates a cGAS immune signaling pathway.

Jessica W Luzwick, Eszter Dombi, Rebecca A Boisvert, Sunetra Roy, Soyoung Park, Selvi Kunnimalaiyaan, Steffi Goffart, Detlev Schindler, Katharina Schlacher.

[Figure: see text].

DOI: https://doi.org/10.1126/sciadv.abf9441
Essays Biochem. 2021 Dec 13. pii: EBC20210036. [Epub ahead of print]

PINK1 signalling in neurodegenerative disease.

Daniel R Whiten, Dezerae Cox, Carolyn M Sue.

  PTEN-induced kinase 1 (PINK1) impacts cell health and human pathology through diverse pathways. The strict processing of full-length PINK1 on the outer mitochondrial membrane populates a cytoplasmic pool of cleaved PINK1 (cPINK1) that is constitutively degraded. However, despite rapid proteasomal clearance, cPINK1 still appears to exert quality control influence over the neuronal protein homeostasis network, including protein synthesis and degradation machineries. The cytoplasmic concentration and activity of this molecule is therefore a powerful sensor that coordinates aspects of mitochondrial and cellular health. In addition, full-length PINK1 is retained on the mitochondrial membrane following depolarisation, where it is a powerful inducer of multiple mitophagic pathways. This function is executed primarily through the phosphorylation of several ubiquitin ligases, including its most widely studied substrate Parkin. Furthermore, the phosphorylation of both pro- and anti-apoptotic proteins by mitochondrial PINK1 acts as a pro-cellular survival signal when faced with apoptotic stimuli. Through these varied roles PINK1 directly influences functions central to cell dysfunction in neurodegenerative disease.

Keywords:  Mitochondria; Mitophagy; Neurodegneration; PTEN induced putative kinase 1; Parkinsons disease

DOI:  https://doi.org/10.1042/EBC20210036
Invest Ophthalmol Vis Sci. 2021 Dec 01. 62(15): 12

SSBP1-Disease Update: Expanding the Genetic and Clinical Spectrum, Reporting Variable Penetrance and Confirming Recessive Inheritance.

Genomics England Research Consortium

Purpose: To report novel genotypes and expand the phenotype spectrum of SSBP1-disease and explore potential disease mechanism.
Methods: Five families with previously unsolved optic atrophy and retinal dystrophy underwent whole genome sequencing as part of the National Institute for Health Research BioResource Rare-Diseases and the UK's 100,000 Genomes Project. In silico analysis and protein modelling was performed on the identified variants. Deep phenotyping including retinal imaging and International Society for Clinical Electrophysiology of Vision standard visual electrophysiology was performed.
Results: Seven individuals from five unrelated families with bilateral optic atrophy and/or retinal dystrophy with extraocular signs and symptoms in some are described. In total, 6 SSBP1 variants were identified including the previously unreported variants: c.151A>G, p.(Lys51Glu), c.335G>A p.(Gly112Glu), and c.380G>A, p.(Arg127Gln). One individual was found to carry biallelic variants (c.380G>A p.(Arg127Gln); c.394A>G p.(Ile132Val)) associated with likely autosomal recessive SSBP1-disease. In silico analysis predicted all variants to be pathogenic and Three-dimensional protein modelling suggested possible disease mechanisms via decreased single-stranded DNA binding affinity or impaired higher structure formation.
Conclusions: SSBP1 is essential for mitochondrial DNA replication and maintenance, with defects leading to a spectrum of disease that includes optic atrophy and/or retinal dystrophy, occurring with or without extraocular features. This study provides evidence of intrafamilial variability and confirms the existence of an autosomal recessive inheritance in SSBP1-disease consequent upon a previously unreported genotype.

DOI: https://doi.org/10.1167/iovs.62.15.12
Am J Med Genet A. 2021 Dec 14.

Rare presentation of FDX2-related disorder and untargeted global metabolomics findings.

Anjali Aggarwal, Nishitha R Pillai, Charles J Billington, Lynn Schema, Susan A Berry.

  We present the case of a 20-year-old male with a history of myopathy and multiple episodes of rhabdomyolysis, and lactic acidosis. He needed hemodialysis for severe rhabdomyolysis-related acute renal failure at the time of initial presentation (age 10 years). Exome sequencing detected a homozygous likely pathogenic variant in FDX2 (c.12G>T, p.M4I). The FDX2 gene encodes a mitochondrial protein, ferredoxin 2, that is involved in the biogenesis of Fe-S clusters. Biallelic pathogenic variants in FDX2 have previously been associated with episodic mitochondrial myopathy with or without optic atrophy and reversible leukoencephalopathy. Only two cases with FDX2-related rhabdomyolysis as a predominant feature have been reported in medical literature. Here, we report a third patient with FDX2-related recurrent, severe episodes of rhabdomyolysis and lactic acidosis. He does not have optic atrophy or leukoencephalopathy. This is the oldest patient reported with FDX2-related disorder and he has significantly elevated CK during episodes of rhabdomyolysis. In addition, we describe untargeted global metabolomic findings during an episode of metabolic decompensation, shedding light on the biochemical pathway perturbation associated with this ultra-rare genetic disorder.

Keywords:  FDX2; metabolomics; rhabdomyolysis

DOI:  https://doi.org/10.1002/ajmg.a.62608
BMC Biol. 2021 Dec 15. 19(1): 265

Enhanced hepatic respiratory capacity and altered lipid metabolism support metabolic homeostasis during short-term hypoxic stress.

Katie A O'Brien, Ben D McNally, Alice P Sowton, Antonio Murgia, James Armitage, Luke W Thomas, Fynn N Krause, Lucas A Maddalena, Ian Francis, Stefan Kavanagh, Dominic P Williams, Margaret Ashcroft, Julian L Griffin, Jonathan J Lyon, Andrew J Murray.

   BACKGROUND: Tissue hypoxia is a key feature of several endemic hepatic diseases, including alcoholic and non-alcoholic fatty liver disease, and organ failure. Hypoxia imposes a severe metabolic challenge on the liver, potentially disrupting its capacity to carry out essential functions including fuel storage and the integration of lipid metabolism at the whole-body level. Mitochondrial respiratory function is understood to be critical in mediating the hepatic hypoxic response, yet the time-dependent nature of this response and the role of the respiratory chain in this remain unclear.
RESULTS: Here, we report that hepatic respiratory capacity is enhanced following short-term exposure to hypoxia (2 days, 10% O2) and is associated with increased abundance of the respiratory chain supercomplex III2+IV and increased cardiolipin levels. Suppression of this enhanced respiratory capacity, achieved via mild inhibition of mitochondrial complex III, disrupted metabolic homeostasis. Hypoxic exposure for 2 days led to accumulation of plasma and hepatic long chain acyl-carnitines. This was observed alongside depletion of hepatic triacylglycerol species with total chain lengths of 39-53 carbons, containing palmitic, palmitoleic, stearic, and oleic acids, which are associated with de novo lipogenesis. The changes to hepatic respiratory capacity and lipid metabolism following 2 days hypoxic exposure were transient, becoming resolved after 14 days in line with systemic acclimation to hypoxia and elevated circulating haemoglobin concentrations.
CONCLUSIONS: The liver maintains metabolic homeostasis in response to shorter term hypoxic exposure through transient enhancement of respiratory chain capacity and alterations to lipid metabolism. These findings may have implications in understanding and treating hepatic pathologies associated with hypoxia.

Keywords:  De novo lipogenesis; Hepatic mitochondria; Hypoxia; Mitochondrial respiratory chain; Mitochondrial supercomplexes

DOI:  https://doi.org/10.1186/s12915-021-01192-0
Cell Death Discov. 2021 Dec 14. 7(1): 389

TRAP1 inhibits MIC60 ubiquitination to mitigate the injury of cardiomyocytes and protect mitochondria in extracellular acidosis.

Lingxiao Zhang, Ning Su, Yuanyuan Luo, Siyin Chen, Tongfeng Zhao.

Extracellular acidosis-induced mitochondrial damage of cardiomyocytes leads to cardiac dysfunction, but no detailed mechanism or efficient therapeutic target has been reported. Here we found that the protein levels of MIC60 were decreased in H9C2 cells and heart tissues in extracellular acidosis, which caused mitochondrial damage and cardiac dysfunction. Overexpression of MIC60 maintains H9C2 cells viability, increases ATP production and mitochondrial membrane potential, mitigates the disruptions of mitochondrial structure and cardiac injury. Mechanistically, extracellular acidosis excessively promoted MIC60 ubiquitin-dependent degradation. TRAP1 mitigated acidosis-induced mitochondrial impairments and cardiac injury by directly interacting with MIC60 to decrease its ubiquitin-dependent degradation in extracellular acidosis.

DOI: https://doi.org/10.1038/s41420-021-00786-5
Hum Mol Genet. 2021 Dec 13. pii: ddab358. [Epub ahead of print]

Genetic landscape of human mitochondrial genome using whole genome sequencing.

Yijing Wang, Guihu Zhao, Zhenghuan Fang, Hongxu Pan, Yuwen Zhao, Yige Wang, Xun Zhou, Xiaomeng Wang, Tengfei Luo, Yi Zhang, Zheng Wang, Qian Chen, Lijie Dong, Yuanfeng Huang, Qiao Zhou, Lu Xia, Bin Li, Jifeng Guo, Kun Xia, Beisha Tang, Jinchen Li.

Increasing evidences suggest that mitochondrial dysfunction is implicated in diseases and aging, and whole-genome sequencing (WGS) is the most unbiased method in analyzing the mitochondrial genome (mtDNA). However, the genetic landscape of mtDNA in the Chinese population has not been fully examined. Here, we described the genetic landscape of mtDNA using WGS data from Chinese individuals (n = 3241). We identified 3892 mtDNA variants, of which 3349 (86%) were rare variants. Interestingly, we observed a trend toward extreme heterogeneity of mtDNA variants. Our study observed a distinct purifying selection on mtDNA, which inhibits the accumulation of harmful heteroplasmies at the individual level: (1) mitochondrial dN/dS ratios were much less than 1; (2) the dN/dS ratio of heteroplasmies was higher than homoplasmies; (3) heteroplasmies had more indels and predicted deleterious variants than homoplasmies. Furthermore, we found that haplogroup M (20.27%) and D (20.15%) had the highest frequencies in the Chinese population, followed by B (18.51%) and F (16.45%). The number of variants per individual differed across haplogroup groups, with a higher number of homoplasmies for the M lineage. Meanwhile, mtDNA copy number was negatively correlated with age but positively correlated with the female sex. Finally, we developed an mtDNA variation database of Chinese populations called MTCards (http://genemed.tech/mtcards/) to facilitate the query of mtDNA variants in this study. In summary, these findings contribute to different aspects of understanding mtDNA, providing a better understanding of the genetic basis of mitochondrial-related diseases.

DOI: https://doi.org/10.1093/hmg/ddab358
PLoS Biol. 2021 Dec;19(12): e3001480

Mutations in LRRK2 linked to Parkinson disease sequester Rab8a to damaged lysosomes and regulate transferrin-mediated iron uptake in microglia.

Adamantios Mamais, Jillian H Kluss, Luis Bonet-Ponce, Natalie Landeck, Rebekah G Langston, Nathan Smith, Alexandra Beilina, Alice Kaganovich, Manik C Ghosh, Laura Pellegrini, Ravindran Kumaran, Ioannis Papazoglou, George R Heaton, Rina Bandopadhyay, Nunziata Maio, Changyoun Kim, Matthew J LaVoie, David C Gershlick, Mark R Cookson.

Mutations in leucine-rich repeat kinase 2 (LRRK2) cause autosomal dominant Parkinson disease (PD), while polymorphic LRRK2 variants are associated with sporadic PD. PD-linked mutations increase LRRK2 kinase activity and induce neurotoxicity in vitro and in vivo. The small GTPase Rab8a is a LRRK2 kinase substrate and is involved in receptor-mediated recycling and endocytic trafficking of transferrin, but the effect of PD-linked LRRK2 mutations on the function of Rab8a is poorly understood. Here, we show that gain-of-function mutations in LRRK2 induce sequestration of endogenous Rab8a to lysosomes in overexpression cell models, while pharmacological inhibition of LRRK2 kinase activity reverses this phenotype. Furthermore, we show that LRRK2 mutations drive association of endocytosed transferrin with Rab8a-positive lysosomes. LRRK2 has been nominated as an integral part of cellular responses downstream of proinflammatory signals and is activated in microglia in postmortem PD tissue. Here, we show that iPSC-derived microglia from patients carrying the most common LRRK2 mutation, G2019S, mistraffic transferrin to lysosomes proximal to the nucleus in proinflammatory conditions. Furthermore, G2019S knock-in mice show a significant increase in iron deposition in microglia following intrastriatal LPS injection compared to wild-type mice, accompanied by striatal accumulation of ferritin. Our data support a role of LRRK2 in modulating iron uptake and storage in response to proinflammatory stimuli in microglia.

DOI: https://doi.org/10.1371/journal.pbio.3001480
World J Hepatol. 2021 Nov 27. 13(11): 1707-1726

Mitochondrial hepatopathy: Respiratory chain disorders- 'breathing in and out of the liver'.

Amrit Gopan, Moinak Sen Sarma.

  Mitochondria, the powerhouse of a cell, are closely linked to the pathophysiology of various common as well as not so uncommon disorders of the liver and beyond. Evolution supports a prokaryotic descent, and, unsurprisingly, the organelle is worthy of being labeled an organism in itself. Since highly metabolically active organs require a continuous feed of energy, any dysfunction in the structure and function of mitochondria can have variable impact, with the worse end of the spectrum producing catastrophic consequences with a multisystem predisposition. Though categorized a hepatopathy, mitochondrial respiratory chain defects are not limited to the liver in time and space. The liver involvement is also variable in clinical presentation as well as in age of onset, from acute liver failure, cholestasis, or chronic liver disease. Other organs like eye, muscle, central and peripheral nervous system, gastrointestinal tract, hematological, endocrine, and renal systems are also variably involved. Diagnosis hinges on recognition of subtle clinical clues, screening metabolic investigations, evaluation of the extra-hepatic involvement, and role of genetics and tissue diagnosis. Treatment is aimed at both circumventing the acute metabolic crisis and long-term management including nutritional rehabilitation. This review lists and discusses the burden of mitochondrial respiratory chain defects, including various settings when to suspect, their evolution with time, including certain specific disorders, their tiered evaluation with diagnostic algorithms, management dilemmas, role of liver transplantation, and the future research tools.

Keywords:  DNA depletion syndrome; Maternal inheritance; Mitochondrial hepatopathy; Neonatal liver failure; Pearson syndrome; Respiratory chain defects

DOI:  https://doi.org/10.4254/wjh.v13.i11.1707
Mitochondrion. 2021 Dec 10. pii: S1567-7249(21)00169-0. [Epub ahead of print]

The molecular role of Sigmar1 in regulating mitochondrial function through mitochondrial localization in cardiomyocytes.

Chowdhury S Abdullah, Richa Aishwarya, Shafiul Alam, Naznin Sultana Remex, Mahboob Morshed, Sadia Nitu, Sumitra Miriyala, Manikandan Panchatcharam, Brandon Hartman, Judy King, Mohammad Alfrad Nobel Bhuiyan, James Traylor, Christopher G Kevil, A Wayne Orr, Shenuarin Bhuiyan.

  Sigmar1 is a widely expressed molecular chaperone protein in mammalian cell systems. Accumulating research demonstrated the cardioprotective roles of pharmacologic Sigmar1 activation by ligands in preclinical rodent models of cardiac injury. Extensive biochemical and immuno-electron microscopic research demonstrated Sigmar1's sub-cellular localization largely depends on cell and organ types. Despite comprehensive studies, Sigmar1's direct molecular role in cardiomyocytes remains elusive. In the present study, we determined Sigmar1's subcellular localization, transmembrane topology, and function using complementary microscopy, biochemical, and functional assays in cardiomyocytes. Quantum dots in transmission electron microscopy showed Sigmar1 labeled quantum dots on the mitochondrial membranes, lysosomes, and sarcoplasmic reticulum-mitochondrial interface. Subcellular fractionation of heart cell lysates confirmed Sigmar1's localization in purified mitochondria fraction and lysosome fraction. Immunocytochemistry confirmed Sigmar1 colocalization with mitochondrial proteins in isolated adult mouse cardiomyocytes. Sigmar1's mitochondrial localization was further confirmed by Sigmar1 colocalization with Mito-Tracker in isolated mouse heart mitochondria. A series of biochemical experiments, including alkaline extraction and proteinase K treatment of purified heart mitochondria, demonstrated Sigmar1 as an integral mitochondrial membrane protein. Sigmar1's structural requirement for mitochondrial localization was determined by expressing FLAG-tagged Sigmar1 fragments in cells. Full-length Sigmar1 and Sigmar1's C terminal-deletion fragments were able to localize to the mitochondrial membrane, whereas N- terminal deletion fragment was unable to incorporate into the mitochondria. Finally, functional assays using extracellular flux analyzer and high-resolution respirometry showed Sigmar1 siRNA knockdown significantly altered mitochondrial respiration in cardiomyocytes. Overall, we found that Sigmar1 localizes to mitochondrial membranes and is indispensable for maintaining mitochondrial respiratory homeostasis in cardiomyocytes.

Keywords:  Sigma-1 receptor; mitochondria; subcellular localization

DOI:  https://doi.org/10.1016/j.mito.2021.12.002
Nat Metab. 2021 Dec 13.

Reducing NAD(H) to amplify rhythms.

Karthikeyani Chellappa, Joseph A Baur.

DOI: https://doi.org/10.1038/s42255-021-00494-5
RMD Open. 2021 Dec;pii: e002010. [Epub ahead of print]7(3):

Circulating mitochondrial DNA copy numbers represent a sensitive marker for diagnosis and monitoring of disease activity in systemic lupus erythematosus.

Stavros Giaglis, Douglas Daoudlarian, Reinhard E Voll, Diego Kyburz, Nils Venhoff, Ulrich A Walker.

   OBJECTIVES: Cell-free DNA is involved in the pathogenesis of systemic lupus erythematosus (SLE) but the clinical value of cell-free DNA measurements in SLE is unknown. Our aim was therefore to examine the utility of mitochondrial (mt) DNA and nuclear (n) DNA quantification in SLE.
METHODS: EDTA plasma was drawn from 103 consecutive patients with SLE and from 56 healthy blood donors. mtDNA and nDNA copy numbers were quantified by PCR from cell-free plasma. Clinical parameters were recorded prospectively.
RESULTS: Circulating mtDNA copy numbers were increased 8.8-fold in the plasma of patients with SLE (median 6.6×107 /mL) compared with controls (median 7.6×106 /mL, p<0.0001). Among all 159 individuals, a cut-off set at 1.8×107 mtDNA copies in a receiver operated curve identified patients with SLE with 87.4% sensitivity and 94.6% specificity; the area under the curve was 0.95 (p<0.0001). mtDNA levels were independent of age or gender, but correlated with Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) on multivariable analysis (p=0.004). Conversely, SLEDAI was associated with prednisone dose (p<0.001), anti-double stranded DNA-titres (p=0.003) and mtDNA levels (p=0.005), but not nDNA copy numbers. In 33 patients with SLE with available follow-up, the changes of mtDNA, but not those of nDNA concentrations, robustly correlated with the evolution of the SLEDAI (r=0.55, p=0.001).
CONCLUSIONS: Circulating mtDNA unlike nDNA molecules are markedly increased in SLE plasma. Regardless of disease activity, circulating mtDNA levels distinguish patients with SLE from healthy controls with high sensitivity and represent an independent marker of SLE activity.

Keywords:  autoimmune diseases; inflammation; lupus erythematosus; systemic

DOI:  https://doi.org/10.1136/rmdopen-2021-002010
Methods Mol Biol. 2021 Dec 15.

Culturing Human Pluripotent Stem Cells on Micropatterned Silicon Surfaces.

Varvara Chalmantzi, Chara Simitzi, Angelos Papadopoulos, Eleni Bagli, Carol Murphy, Emmanuel Stratakis, Theodore Fotsis.

  Human pluripotent stem cell culture conditions are constantly being optimized, thus providing insight to the environmental cues that affect cell choices. A wide variety of media, coating materials, and substrates is now available for use, serving different scientific needs. Factors such as material stiffness, roughness, and topography are being recognized to contribute or even direct the acquisition of specific phenotypes. Here, we describe the use of patterned silicon substrates coated with Matrigel for the propagation and differentiation of human pluripotent stem cells.

Keywords:  BRACHYURY; Immunofluorescence; Laser manufacturing; NANOG; Silicon substrates; Surface micropatterning; Topography; hPSCs

DOI:  https://doi.org/10.1007/7651_2021_428
Nat Metab. 2021 Dec 13.

What is the most appropriate covariate in ANCOVA when analysing metabolic rate?

Sam Virtue, Christopher J Lelliott, Antonio Vidal-Puig.

DOI: https://doi.org/10.1038/s42255-021-00505-5
Front Physiol. 2021 ;12 718982

Neurogenic Hypertension Mediated Mitochondrial Abnormality Leads to Cardiomyopathy: Contribution of UPRmt and Norepinephrine-miR- 18a-5p-HIF-1α Axis.

Shyam S Nandi, Kenichi Katsurada, Sushil K Mahata, Kaushik P Patel.

  Aims: Hypertension increases the risk of heart disease. Hallmark features of hypertensive heart disease is sympathoexcitation and cardiac mitochondrial abnormality. However, the molecular mechanisms for specifically neurally mediated mitochondrial abnormality and subsequent cardiac dysfunction are unclear. We hypothesized that enhanced sympatho-excitation to the heart elicits cardiac miR-18a-5p/HIF-1α and mitochondrial unfolded protein response (UPRmt) signaling that lead to mitochondrial abnormalities and consequent pathological cardiac remodeling. Methods and Results: Using a model of neurogenic hypertension (NG-HTN), induced by intracerebroventricular (ICV) infusion of Ang II (NG-HTN; 20 ng/min, 14 days, 0.5 μl/h, or Saline; Control, 0.9%) through osmotic mini-pumps in Sprague-Dawley rats (250-300 g), we attempted to identify a link between sympathoexcitation (norepinephrine; NE), miRNA and HIF-1α signaling and UPRmt to produce mitochondrial abnormalities resulting in cardiomyopathy. Cardiac remodeling, mitochondrial abnormality, and miRNA/HIF-1α signaling were assessed using histology, immunocytochemistry, electron microscopy, Western blotting or RT-qPCR. NG-HTN demonstrated increased sympatho-excitation with concomitant reduction in UPRmt, miRNA-18a-5p and increased level of HIF-1α in the heart. Our in silico analysis indicated that miR-18a-5p targets HIF-1α. Direct effects of NE on miRNA/HIF-1α signaling and mitochondrial abnormality examined using H9c2 rat cardiomyocytes showed NE reduces miR-18a-5p but increases HIF-1α. Electron microscopy revealed cardiac mitochondrial abnormality in NG-HTN, linked with hypertrophic cardiomyopathy and fibrosis. Mitochondrial unfolded protein response was decreased in NG-HTN indicating mitochondrial proteinopathy and proteotoxic stress, associated with increased mito-ROS and decreased mitochondrial membrane potential (ΔΨm), and oxidative phosphorylation. Further, there was reduced cardiac mitochondrial biogenesis and fusion, but increased mitochondrial fission, coupled with mitochondrial impaired TIM-TOM transport and UPRmt. Direct effects of NE on H9c2 rat cardiomyocytes also showed cardiomyocyte hypertrophy, increased mitochondrial ROS generation, and UPRmt corroborating the in vivo data. Conclusion: In conclusion, enhanced sympatho-excitation suppress miR-18a-5p/HIF-1α signaling and increased mitochondrial stress proteotoxicity, decreased UPRmt leading to decreased mitochondrial dynamics/OXPHOS/ΔΨm and ROS generation. Taken together, these results suggest that ROS induced mitochondrial transition pore opening activates pro-hypertrophy/fibrosis/inflammatory factors that induce pathological cardiac hypertrophy and fibrosis commonly observed in NG-HTN.

Keywords:  HIF-1α; UPRmt; cardiac fibrosis; cardiac hypertrophy; hypertension; miR-18a-5p; mitochondrial damage

DOI:  https://doi.org/10.3389/fphys.2021.718982
BMB Rep. 2021 Dec 14. pii: 5459. [Epub ahead of print]

Reactive microglia and mitochondrial unfolded protein response following ventriculomegaly and behavior defects in kaolin-induced hydrocephalus.

Jiebo Zhu, Min Joung Lee, Hee Jin Chang, Xianshu Ju, Jianchen Cui, Yu Lim Lee, Dahyun Go, Woosuk Chung, Eungseok Oh, Jun Young Heo.

Ventriculomegaly induced by the abnormal accumulation of cerebrospinal fluid (CSF) leads to hydrocephalus, which is accompanied by neuroinflammation and mitochondrial oxidative stress. The mitochondrial stress activates mitochondrial unfolded protein response (UPRmt), which is essential for mitochondrial protein homeostasis. However, the association of inflammatory response and UPRmt in the pathogenesis of hydrocephalus is still unclear. To assess their relevance in the pathogenesis of hydrocephalus, we established a kaolin-induced hydrocephalus model in 8-week-old male C57BL/6J mice and evaluated it over time. We found that kaolin-injected mice showed prominent ventricular dilation, motor behavior defects at the 3-day, followed by the activation of microglia and UPRmt in the motor cortex at the 5-day. In addition, PARP-1/NF-κB signaling and apoptotic cell death appeared at the 5-day. Taken together, our findings demonstrate that activation of microglia and UPRmt occurs after hydrocephalic ventricular expansion and behavioral abnormalities which could be lead to apoptotic neuronal cell death, providing a new perspective on the pathogenic mechanism of hydrocephalus.
Methods Mol Biol. 2022 ;2420 159-175

Development of a Standardized MRM Method for the Quantification of One Carbon Metabolism Enzymes.

Laura Guerrero, Alberto Paradela, Fernando J Corrales.

  One-carbon metabolism (1CM) plays a central role in liver physiology, being the source of essential metabolites such as S-adenosylmethionine, the main alkylating agent in living cells, and glutathione, their most important nonenzymatic antioxidant defense. Impairment of 1CM in hepatocytes is a recognized factor associated to chronic liver disorders and hepatocellular carcinoma. With this in mind, we have proposed the concept of functional biomarker referring to a cellular pathway that can be systematically monitored as indicative of a particular physiological or pathological condition. Here we describe a targeted mass spectrometry (MRM) protocol to simultaneously quantify 13 1CM enzymes in liver tissue specimens.

Keywords:  Hepatocellular carcinoma; Liver diseases; Mass spectrometry; Multiple reaction monitoring; One-carbon metabolism; Targeted proteomics

DOI:  https://doi.org/10.1007/978-1-0716-1936-0_13
iScience. 2021 Dec 17. 24(12): 103438

Murine cerebral organoids develop network of functional neurons and hippocampal brain region identity.

Francesca Ciarpella, Raluca Georgiana Zamfir, Alessandra Campanelli, Elisa Ren, Giulia Pedrotti, Emanuela Bottani, Andrea Borioli, Davide Caron, Marzia Di Chio, Sissi Dolci, Annika Ahtiainen, Giorgio Malpeli, Giovanni Malerba, Rita Bardoni, Guido Fumagalli, Jari Hyttinen, Francesco Bifari, Gemma Palazzolo, Gabriella Panuccio, Giulia Curia, Ilaria Decimo.

  Brain organoids are in vitro three-dimensional (3D) self-organized neural structures, which can enable disease modeling and drug screening. However, their use for standardized large-scale drug screening studies is limited by their high batch-to-batch variability, long differentiation time (10-20 weeks), and high production costs. This is particularly relevant when brain organoids are obtained from human induced pluripotent stem cells (iPSCs). Here, we developed, for the first time, a highly standardized, reproducible, and fast (5 weeks) murine brain organoid model starting from embryonic neural stem cells. We obtained brain organoids, which progressively differentiated and self-organized into 3D networks of functional neurons with dorsal forebrain phenotype. Furthermore, by adding the morphogen WNT3a, we generated brain organoids with specific hippocampal region identity. Overall, our results showed the establishment of a fast, robust and reproducible murine 3D in vitro brain model that may represent a useful tool for high-throughput drug screening and disease modeling.

Keywords:  Biological sciences; Cell biology; Developmental biology; Neuroscience

DOI:  https://doi.org/10.1016/j.isci.2021.103438
Mitochondrion. 2021 Dec 13. pii: S1567-7249(21)00170-7. [Epub ahead of print]

Altering neuronal circuitry with 4-aminopyridine for visual improvement in leber's hereditary optic neuropathy (LHON).

Jane W Chan, William Sultan, Rustum Karanjia, Alfredo A Sadun.

  In this retrospective, interventional, longitudinal small case series, we looked at the visual effects of pharmacologic intervention with 4-aminopyridine (4-AP) in chronic Leber's Hereditary Optic Neuropathy (LHON) patients who are non-responders to idebenone. We illustrate, as examples, the visual progression of three LHON patients with 4-AP as add-on therapy to idebenone. Each patient had a different primary LHON mutation and was treated with idebenone within one year of onset. No response to idebenone at 300 mg orally three times a day ranged from less than one year to 2.5 years, and the addition of 4-AP at 10 mg orally two times a day ranged from 24 to 29 months. Outcome measures included best-corrected distance visual acuity, color vision, automated perimetry, the average retinal nerve fiber layer (RNFL) thickness, and the full-field photopic negative response (PhNR) amplitude. The 19-year-old man with the LHON mutation 11778A>G had no response to the addition of 4-AP to idebenone. The 27-year-old man with the LHON mutation 3460A>G experienced a significant response to 4-AP. Finally, the 40-year-old man with the LHON mutation 14484T>C had a milder response. Although this case series was too small to demonstrate the efficacy of idebenone with add-on 4AP, it allowed us to consider a new hypothesis that neuronal activity generated from 4-AP can add more potential for visual recovery in LHON patients.

Keywords:  4-aminopyridine; Idebenone; Leber’s hereditary optic neuropathy (LHON); Neuroenhancement; Neuronal activity

DOI:  https://doi.org/10.1016/j.mito.2021.12.003
Genet Med. 2021 Nov 17. pii: S1098-3600(21)01121-7. [Epub ahead of print]

DNA methylation episignature testing improves molecular diagnosis of Mendelian chromatinopathies.

Jennifer Kerkhof, Gabriella Maria Squeo, Haley McConkey, Michael A Levy, Maria Rosaria Piemontese, Marco Castori, Maria Accadia, Elisa Biamino, Matteo Della Monica, Marilena Carmela Di Giacomo, Cristina Gervasini, Silvia Maitz, Daniela Melis, Donatella Milani, Maria Piccione, Paolo Prontera, Angelo Selicorni, Bekim Sadikovic, Giuseppe Merla.

   PURPOSE: Chromatinopathies include more than 50 disorders caused by disease-causing variants of various components of chromatin structure and function. Many of these disorders exhibit unique genome-wide DNA methylation profiles, known as episignatures. In this study, the methylation profile of a large cohort of individuals with chromatinopathies was analyzed for episignature detection.
METHODS: DNA methylation data was generated on extracted blood samples from 129 affected individuals with the Illumina Infinium EPIC arrays and analyzed using an established bioinformatic pipeline.
RESULTS: The DNA methylation profiles matched and confirmed the sequence findings in both the discovery and validation cohorts. Twenty-five affected individuals carrying a variant of uncertain significance, did not show a methylation profile matching any of the known episignatures. Three additional variant of uncertain significance cases with an identified KDM6A variant were re-classified as likely pathogenic (n = 2) or re-assigned as Wolf-Hirschhorn syndrome (n = 1). Thirty of the 33 Next Generation Sequencing negative cases did not match a defined episignature while three matched Kabuki syndrome, Rubinstein-Taybi syndrome and BAFopathy respectively.
CONCLUSION: With the expanding clinical utility of the EpiSign assay, DNA methylation analysis should be considered part of the testing cascade for individuals presenting with clinical features of Mendelian chromatinopathy disorders.

Keywords:  Chromatinopathies; DNA methylation; Epigenetics

DOI:  https://doi.org/10.1016/j.gim.2021.08.007
Genet Med. 2021 Nov 18. pii: S1098-3600(21)04743-2. [Epub ahead of print]

The importance of automation in genetic diagnosis: Lessons from analyzing an inherited retinal degeneration cohort with the Mendelian Analysis Toolkit (MATK).

Erin Zampaglione, Matthew Maher, Emily M Place, Naomi E Wagner, Stephanie DiTroia, Katherine R Chao, Eleina England, Broad Cmg, Andrew Catomeris, Sherwin Nassiri, Seraphim Himes, Joey Pagliarulo, Charles Ferguson, Eglé Galdikaité-Braziené, Brian Cole, Eric A Pierce, Kinga M Bujakowska.

   PURPOSE: In Mendelian disease diagnosis, variant analysis is a repetitive, error-prone, and time consuming process. To address this, we have developed the Mendelian Analysis Toolkit (MATK), a configurable, automated variant ranking program.
METHODS: MATK aggregates variant information from multiple annotation sources and uses expert-designed rules with parameterized weights to produce a ranked list of potentially causal solutions. MATK performance was measured by a comparison between MATK-aided and human-domain expert analyses of 1060 families with inherited retinal degeneration (IRD), analyzed using an IRD-specific gene panel (589 individuals) and exome sequencing (471 families).
RESULTS: When comparing MATK-assisted analysis with expert curation in both the IRD-specific gene panel and exome sequencing (1060 subjects), 97.3% of potential solutions found by experts were also identified by the MATK-assisted analysis (541 solutions identified with MATK of 556 solutions found by conventional analysis). Furthermore, MATK-assisted analysis identified 114 additional potential solutions from the 504 cases unsolved by conventional analysis.
CONCLUSION: MATK expedites the process of identification of likely solving variants in Mendelian traits, and reduces variability stemming from human error and researcher bias. MATK facilitates data reanalysis to keep up with the constantly improving annotation sources and next-generation sequencing processing pipelines. The software is open source and available at https://gitlab.com/matthew_maher/mendelanalysis.

Keywords:  Automation; Mendelian analysis; Variant ranking

DOI:  https://doi.org/10.1016/j.gim.2021.09.015
Cell Rep. 2021 Dec 14. pii: S2211-1247(21)01602-8. [Epub ahead of print]37(11): 110108

Sarm1 haploinsufficiency or low expression levels after antisense oligonucleotides delay programmed axon degeneration.

Stacey Anne Gould, Jonathan Gilley, Karen Ling, Paymaan Jafar-Nejad, Frank Rigo, Michael Coleman.

  Activation of the pro-degenerative protein SARM1 after diverse physical and disease-relevant injuries causes programmed axon degeneration. Original studies indicate that substantially decreased SARM1 levels are required for neuroprotection. However, we demonstrate, in Sarm1 haploinsufficient mice, that lowering SARM1 levels by 50% delays programmed axon degeneration in vivo after sciatic nerve transection and partially prevents neurite outgrowth defects in mice lacking the pro-survival factor NMNAT2. In vitro, the rate of degeneration in response to traumatic, neurotoxic, and genetic triggers of SARM1 activation is also slowed. Finally, we demonstrate that Sarm1 antisense oligonucleotides decrease SARM1 levels by more than 50% in vitro, which delays or prevents programmed axon degeneration. Combining Sarm1 haploinsufficiency with antisense oligonucleotides further decreases SARM1 levels and prolongs protection after neurotoxic injury. These data demonstrate that axon protection occurs in a Sarm1 gene dose-responsive manner and that SARM1-lowering agents have therapeutic potential, making Sarm1-targeting antisense oligonucleotides a promising therapeutic strategy.

Keywords:  ALS; NMNAT2; SARM1; Wallerian degeneration; antisense oligonucleotides; neurodegeneration; neuroprotection; peripheral neuropathy; programmed axon degeneration; therapy

DOI:  https://doi.org/10.1016/j.celrep.2021.110108
Genet Med. 2021 Nov 17. pii: S1098-3600(21)01129-1. [Epub ahead of print]

Cost-effectiveness of genome sequencing for diagnosing patients with undiagnosed rare genetic diseases.

Devin Incerti, Xiang-Ming Xu, Jacquelyn W Chou, Nina Gonzaludo, John W Belmont, Brock E Schroeder.

   PURPOSE: To estimate the cost-effectiveness of genome sequencing (GS) for diagnosing critically ill infants and noncritically ill pediatric patients (children) with suspected rare genetic diseases from a United States health sector perspective.
METHODS: A decision-analytic model was developed to simulate the diagnostic trajectory of patients. Parameter estimates were derived from a targeted literature review and meta-analysis. The model simulated clinical and economic outcomes associated with 3 diagnostic pathways: (1) standard diagnostic care, (2) GS, and (3) standard diagnostic care followed by GS.
RESULTS: For children, costs of GS ($7284) were similar to that of standard care ($7355) and lower than that of standard care followed by GS pathways ($12,030). In critically ill infants, when cost estimates were based on the length of stay in the neonatal intensive care unit, the lowest cost pathway was GS ($209,472). When only diagnostic test costs were included, the cost per diagnosis was $17,940 for standard, $17,019 for GS, and $20,255 for standard care followed by GS.
CONCLUSION: The results of this economic model suggest that GS may be cost neutral or possibly cost saving as a first line diagnostic tool for children and critically ill infants.

Keywords:  Cost-effectiveness; Genetic disease; Genome sequencing; Rare disease

DOI:  https://doi.org/10.1016/j.gim.2021.08.015